The Research Plan can be divided into five sections. 1. Secondary metal binding sites of sero- and lactotransferrin. Evidence from amino acid sequence comparisons, EPR, kinetics and spectrophotometry reveal the presence of secondary metal binding sites. I wish to ascertain their stoichiometry, relative affinities for different metals, metal exchange reactivities and their effect on the primary metal/anion sites. 2. Iron exchange reactivities of lactotransferrin. Little is known about the reactions in which lactotransferrin attacks, sequesters and releases iron. This information will shed light on possible physiological roles of lactotransferrin and provide a valuable comparison with the data on serotransferrin. 3. Environmental and chemical modification and the iron exchange reactivities of the transferrins. Denaturants, pH, chemical modification, temperature and salts have a differential effect on the sites in the transferrins' two lobes. I want to know how the modification alters the attack, sequestration and iron release properties. Of special interest will be insight into the conformational change that results in the labilization of the iron and anion binding stress. 4. Control of attack, sequestration and release of iron by the anion binding site. The anion binding site controls the affinity of the protein for iron and other metals. This phenomenon can be explored via the use of carbonate substitutes in studies of metal ion attack, sequestration, oxidation, reduction and release. 5. Exchange of metals other than iron by the transferrins. The transferrins offer an excellent model system for examining protein metal ion exchange for Cu, Mn, Co, Cr, Ni and others. I want to discover the rules that govern exchange of metal ions and to learn about the mechanisms that direct metals to specific sites. 6. Metal ion exchange between the transferrins and other proteins. How are metal ions passed from one protein to another? Is it always via mediators such as chelators and redox reagents? or via a modification force such as acidification? Do protein-metal-protein complexes form? Ceruloplasmin, phosvitin, the sero-, ovo- and lactotransferrins, serum albumin, carboxypeptidase A, ferritin will be subjects of study. 7. Lactotransferrin in milk and serotransferrin in serum. Too little attention is paid to the metal ion exchange reactivities in the natural environment. I wish to extend much of the work above to the physiological media of interest.